MicroRNA changes in chronic lymphocytic leukemia in response to ibrutinib treatment. In 2015-2016 we finalized and published our study reporting microRNA levels in lymph node and peripheral blood derived CLL cells from patients pre and post treatment with the BTK inhibitor ibrutinib. This work was done in collaboration with Dr. Adrian Wiestner in NHLBI. We reported 1) unique miR profiles in CLL cells in the lymph node related to B-cell activation that become down regulated upon release into the peripheral blood; 2) a subset of 4 miRNA related to BCR activation become significantly down regulated in circulating CLL cells after treatment with ibrutinib, 3) multiple predicted targets of the 4 miRNA include tumor suppressor transcripts that are significantly upregulated after ibrutinib treatment, and 4) functional studies demonstrating direct regulation of the tumor suppressors by the miRNA. These findings suggest that response to Ibrutinib involves upregulation of tumor suppressors via downregulation of miRNAs, representing a novel mechanism of treatment response. This work was the subject of a platform presentation at the Annual Society of Hematology meeting in December 2015 and was published in Leukemia in 2016 (Saleh L. et al.). MicroRNA profiling of GATA2 deficiency associated myelodysplasia (MDS) / acute myeloid leukemia (AML). Somatic and inherited germline mutations in GATA2 have been identified in patients diagnosed with monocytopenia, B-cell and NK-cell lymphopenia, susceptibility to opportunistic infections (e.g. disseminated MAC), and a strong propensity to develop hypocellular MDS/AML or CMML. We previously identified significantly increased levels of miR-181c in EBV transformed cell lines from patients with GATA2 deficiency. mRNA targets of miR-181c include MCL-1 transcripts involved in the regulation of apoptosis. We performed functional studies validating the regulation of MCL-1 by miR-181c. During 2015-2016 we additionally identified miRNA changes related to markedly increased FLT3 ligand expression in GATA2 and are performing functional studies to validate the relationship between miR-424 and FLT3L. These findings suggest potential mechanisms for the progressive cytopenias in GATA2 deficiency involving down regulation of MCL-1 in stem cells and upregulation of FLT3L in the bone marrow microenvironment. RAS-associated Autoimmune Leukoproliferative Disorder (RALD) is a chronic indolent condition that typically presents in childhood with monocytosis, lymphocytosis, autoimmune phenomena, splenomegaly, and variable lymphadenopathy. RALD is characterized by somatic mutations in KRAS or NRAS in hematopoietic cells. Despite the indolent course of RALD, it shares some overlapping clinical features and similar genetic defects with Juvenile Myelomonocytic Leukemia (JMML), which is characterized by canonical and non-canonical RAS-pathway mutations. The nature of RALD is somewhat controversial as many pediatric hematologist/oncologists are concerned that RALD may represent a form of JMML or a precursor disease with the potential of transforming into JMML. In 2015-2016, we performed targeted sequencing for 54 genes commonly associated with myeloid neoplasia. Mutations in NRAS or KRAS were the only mutations identified suggesting that the most frequent additional cooperating mutations found in JMML, AML, CMML and MDS are not present in RALD. Ongoing studies are in process. Immunophenotypic and morphologic analyses of other rare diseases with bone marrow and peripheral blood pathology. Multiple collaborative research studies continued in 2015-2016 involving immunophenotypic and morphologic characterization of PASLI disease and abnormal B-cell maturation related to mutations in PI3KCD, immunodeficiency and cytopenias related to mutations in CTLA4, B-cell deficiency in patients with mutations involving IKAROS, CANDLE syndrome, aplastic anemia, hypocellular MDS and AML. Our manuscript detailing abnormal B-cell maturation in the bone marrow of patients with germline mutations in PIK3CD was accepted to the Journal of Allergy and Clinical Immunology and is currently in press.